ED Sciences de la Vie et de la Santé
Mitochondrial RNA Processing And Translation In Eukaryotic Parasites
by Sramona BARUA (Acides nucléiques : Régulations Naturelles et Artificielles)
The defense will take place at h00 - Auditorium European Institute of Chemistry and Biology, 2 Rue Robert Escarpit 33607 PESSAC FRANCE
in front of the jury composed of
- Yaser HASHEM - Directeur de recherche - University of Hamburg - Directeur de these
- Petya KRASTEVA - Directrice de recherche - UMR5248 CBMN - Examinateur
- Pascal AUFFINGER - Directeur de recherche - Institute for Molecular and Cellular Biology (IBMC), French National Centre for Scientific Research - Rapporteur
- Eva KOWALINSKI - Directrice de recherche - European Molecular Biology Laboratory (EMBL) — Grenoble Site - Rapporteur
Mitochondria are essential organelles that sustain eukaryotic life by driving energy production and numerous metabolic processes. In parasitic eukaryotes, however, mitochondrial gene expression has evolved in highly divergent ways, reflecting adaptation to complex life cycles and host environments. Central to this divergence are mitochondrial RNA processing and translation, which govern the maturation and decoding of transcripts into functional proteins required for oxidative phosphorylation and parasite survival. Unlike well-studied model organisms, parasitic mitochondria often display unusual genome organization, non-canonical RNA processing pathways, and specialized mitoribosomes. Understanding how mitochondrial RNAs are processed and translated in these organisms is therefore crucial for uncovering fundamental aspects of mitochondrial biology and for identifying parasite-specific vulnerabilities that may be exploited therapeutically. In kinetoplastid parasites, mitochondrial gene expression is distinguished by an extensive RNA processing pathway known as RNA editing. Most mitochondrial transcripts are synthesized as cryptic pre-mRNAs that require post-transcriptional insertion and deletion of uridine residues to become translatable. This process is catalyzed by a large multiprotein complex termed the editosome, which acts together with small guide RNAs that specify editing sites. The editosome carries out a precisely ordered series of reactions involving endonucleolytic cleavage, uridine insertion or removal, and RNA ligation, thereby converting immature transcripts into functional mRNAs essential for parasite survival. Editosomes are indispensable for kinetoplastid viability, and understanding their structure is critical for defining how subunits assemble into a dynamic macromolecular machine, how catalytic and accessory factors are organized, and how substrate recognition and reaction coordination are achieved at the molecular level. Because editosomes are unique to trypanosomatid parasites and absent in mammalian hosts, structural insights can reveal parasite-specific interaction surfaces that offer promising opportunities for selective drug design. Accordingly, this study focuses on the purification and structural characterization of the ~20S editosome complex. By introducing affinity tags into selected subunits, the complex was isolated and analyzed by cryo-electron microscopy to define its molecular architecture and organization, providing a global structural framework for understanding editosome assembly and function at high resolution. While RNA editing represents a remarkable layer of post-transcriptional control in kinetoplastids, mitochondrial gene expression ultimately culminates in translation. In other parasitic lineages such as apicomplexans, mitochondrial RNAs are not extensively edited, yet they are decoded by highly divergent mitochondrial ribosomes. These mitoribosomes have evolved protein-rich architectures to accommodate reduced genomes and atypical transcripts. In the next part of this thesis, I focus on the apicomplexan parasite Toxoplasma gondii, for which I analyzed the cryo-EM structure of the mitochondrial ribosome and its key structural features. We determined and published the first cryo-EM structure of an apicomplexan mitoribosome, reaching resolutions of 2.89 Å for the LSU, 3.6 Å for the SSU head, and 3.2 Å for the SSU body. Structural analysis revealed the most fragmented mitoribosomal rRNA described to date, with 25 fragments in the LSU and 15 in the SSU assigned, and an additional 23 fragments remaining unassigned across both subunits.